Project Summary/Abstract While outcomes have substantially improved for many types of cancer, endometrial cancer incidences and deaths are on the rise, with the five year survival rate worse today than three decades ago; owing largely to the ineffectiveness of current treatments. As a tumor is exquisitely sensitive to the growth promoting effects of estrogen and the growth limiting effects of progesterone, hormonal therapy for endometrial cancer using progestins has been a traditional choice for treatment. It is highly effective in the short term; however, responsiveness wanes over time due to loss of progesterone receptor (PR) expression, and recurrences are common. There is a critical need to identify strategies to improve or restore responsiveness to progestin therapy, and we propose that molecularly enhanced progestin therapy will make a major positive impact on survival of patients with endometrial cancer. The objective of this application is to identify the molecular mechanisms driving the downregulation of the PR in endometrial cancer patients and identify novel strategies to further enhance the effectiveness of progestin therapy. We will develop molecular agent combinations with progestins that will significantly enhance tumor cell differentiation in vitro and improve survival in mouse xenograft models of human endometrial cancer. Our central hypothesis is that targeting PR repressors will enhance the expression of PR, the most important tumor suppressor in the endometrium, thereby improving response to progestin therapy. This hypothesis stems from our strong recently published data in endometrial cancer cells that PR expression is downregulated through distinct molecular mechanisms, and epigenetic modulators potently increase PR expression and tumor suppressor activity. We have now identified additional PR suppressors that have the potential to more clearly define the multiple mechanisms of PR inhibition in endometrial cancer, setting the stage for new therapeutic opportunities. In Aim 1, we will determine the impact of epigenetic modulators on PR expression and activity in endometrial cancer patients from our related clinical trial NRG-GY011 results; in Aim 2, we will enhance PR expression using small molecular drugs and test drug efficacy using in vitro and in vivo endometrial cancer models. In Aim 3, we will identify novel PR downregulation mechanisms using genome-wide gene silencing. At the completion of these studies, it is our expectation that we will have identified mechanisms of hormonal resistance, successfully integrated innovative molecular therapies into enhanced progestin therapeutic regimens in preclinical and clinical studies, and inform for the design of future endometrial cancer clinical trials. As such, these studies have a strong potential to impact the alarming trend towards declining survival in endometrial cancer.